DESCRIPTION (Taken from the application): Osteoporosis is a group of diseases of diverse etiology in which the rate of bone resorption by osteoclasts exceeds that of bone formation by osteoblasts, resulting in a reduction in the mass of bone per unit volume. Patients with end-stage, 1ow-turnover osteoporosis do not respond to existing therapies. Osteoblasts in end-stage osteoporosis can be activated by parathyroid hormone (PTH); however, this also invokes an increase in osteoclastic activity that limits the effectiveness of this approach. In this project, we propose to develop a gene therapy method to allow PTH activation of osteoblasts that will have reduced ability to produce RANKL, a key regulator of osteoclastogenesis. We hypothesize that this could be accomplished by employing an intracellular single-chain antibody (sFv) to achieve osteoblast-specific downregulation of RANKL. This approach mandates the use of a gene delivery vehicle, or vector, which can target expression of the sFv specifically to osteoblasts. Capitalizing on the ability of recombinant adenoviral (Ad) vectors to accomplish efficient gene transfer, we hypothesize that Ad-mediated gene delivery can be targeted specifically to osteoblasts. The first Specific Aim is to develop an Ad vector targeted to osteoblasts at the transductional and transcriptional levels. Transductional targeting will be achieved using a bispecific antibody with specificities for the adenovirus fiber protein and an osteoblastrelated marker protein, while transcriptional targeting will be achieved by placing the transgene under the control of the osteoblast-specific osteocalcin promoter. The second Specific Aim is to derive an intracellular sFv to selectively downregulate the expression of RANKL. In the third Specific Aim, we will employ the targeted Ad vector to direct the intracellular expression of this sFv specifically within osteoblasts and will determine whether the downregulation of RANKL results in a reduction in osteoclast recruitment in vitro. The realization of these Specific Aims would establish the feasibility of this approach as a rational strategy for gene therapy in patients with end-stage, low-turnover osteoporosis.